Staphylococcus aureus is both a common skin colonizer and also a formidable pathogen, causing the majority of skin and soft tissue and surgical site infections in the United States. S. aureus skin infections serve as a site from which bacteria can disseminate to virtually any organ. These invasive infections cause an estimated 11,000 deaths in the U.S. annually. In addition, antibiotic- resistant strains (MRSA) are especially difficult and costly to treat and constitute an escalating public health threat. Understanding how S. aureus persists in and breaches the skin epithelium has the potential to deliver tremendous preventative and therapeutic value. Significant gaps in our knowledge of this important pathogen are understanding how S. aureus virulence factors modify the host epithelial niche for long-term tissue colonization and how S. aureus breaches epithelial barriers during the transition from colonization to infection. The secreted pore- forming virulence factor a-toxin targets the cell-cell junction protein E-cadherin for cleavage and is essential for S. aureus pathogenesis during epithelial infections. Blocking a-toxin activity at the epithelial cell-cell junctions may ultimately be an effective strategy to interfere with MRSA skin colonization and subsequent infection into deeper tissues. With this idea in mind, we performed a genetic screen for novel host factors required for a-toxin virulence by intoxicating mutagenized haploid human cells. Our screen identified a set of genes that encode for members of the epithelial adherens junctions which, when absent, confer resistance to a- toxin. We have validated the most prominent adherens junction gene discovered by our screen for its role in facilitating a-toxin cytotoxicity. We hypothesize that altering expression of this gene will reduce a-toxin virulence in the skin and abrogate morbidity from MRSA infection. In this proposal, we will use in vitro human cell lines, a 3D human epidermal organotypic skin culture model, and a transgenic mouse to study the consequences of a-toxin virulence mediated by the cellular adherens junctions. This F31 training award will provide the experimental resources, time, and training support necessary to identify how the adherens junctions control MRSA pathogenicity. Results from these experiments will likely lead to better strategies to prevent and treat MRSA infections, a national priority for disease control.